Type 1 diabetes (T1D) is an autoimmune disease characterized by T cell-mediated destruction of the pancreatic islet ( cells. Studies of the nonobese diabetic (NOD) mouse model of the disease indicate that CD8+ T cells are required for T1D development. We used ( cell-cytotoxic CD8+ T cell clones isolated from NOD mice to identify two antigenic targets for early insulitic T cells. These are islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) and dystrophia myotonica kinase (DMK). Considered together with the third known target of early insulitic CD8+ T cells (i.e., insulin), these three specificities can collectively account for up to half of the islet-infiltrating CD8+ T cells in NOD mice. Here we will translate our findings to human T1D by using islet-infiltrating T cells from HLA-transgenic NOD mice to map the epitopes of these ( cell antigens that are targeted by HLA-A*0201-restricted T cells. The hypothesis that HLA-A*0201 -positive patients and HLA-transgenic mice will exhibit overlapping specificities with respect to the epitopes they recognize will then be evaluated. Next, our newly developed ability to deliver ( cell antigens to steady-state dendritic cells (DCs) via the endocytic receptor DEC-205 will permit us to test the hypothesis that this treatment will result in CD8+ T cell tolerance and improvement of disease. We will examine the impact of DC targeting of an IGRP-derived peptide on both transferred and endogenous IGRP-reactive CD8+ T cells, utilizing approaches that will enable us to monitor IGRP-specific T cells in terms of both number and function, including a cytotoxicity assay that we have devised to permit detection of IGRP-specific T cell cytotoxicity in vivo. IGRP has been chosen for these studies, because it is a CD8+ T cell antigen of major importance in NOD mice. Finally, a "high-throughput" approach to antigen discovery will be employed to permit a more complete characterization of the antigens targeted by CD8+ T cells in the NOD mouse model, which will increase its utility in optimizing therapies and exploring the immunopathogenesis of T1D. Retroviral transduction of bone marrow stem cells will be used to rapidly generate NOD mice expressing T cell receptors from early insulitic ( cell-cytotoxic CD8+ T cell clones of unknown specificity. These mice will be used as a T cell source for antigen discovery. In summary, our work will facilitate the development of strategies to interfere with pathogenic T cell populations, as well as assays to monitor autoimmune activity. Relevance in lay language: ( cells in the pancreas make insulin, which is essential for controlling how sugar is used in the body. Type 1 diabetes occurs when the immune system kills the ( cells and insulin can no longer be made. The proposed work will identify the molecular targets of the destructive immune cells and develop methods to monitor and control their activity.